Novel alkylated pregnadienes



United States Patent Ofiice 3,013,033 Patented Doc. 12, 1961 3,013,033 NOVEL ALKYLATED PREGNADIENES Alexander L. Nussbaum, Leonia, and Eugene P. Oliveto, Bloomfield, N.J., assignors to Schering Corporation, Bloomfield, N.J., a corporation of New Jersey N Drawing. Filed Oct. 29, 1958, Ser. No. 770,315 9 Claims. (Cl. zen-$97.45

This invention pertains to a new group of therapeutically active pregnadienes and to methods for their manufacture. In particular, our invention relates to 1,5-pregnadiene-l7a,2l-diol-3,20-diones which possess a 'l6-lower alkyl substituent in the aor B-configuration. These novel compounds are potent anti-inflammatory agents.

Our novel compounds may be represented by the f0llowing formula:

wherein X is a member of the group consisting of hydrogen and halogen having an atomic weight less than 126; Y is a member of the group consisting of O and (H, SOH); Z is an alkyl radical preferably having from 1 to 4 carbon atoms; and R is a member of the group consisting of H and acyl.

Illustrative of the 21-esters contemplated above are alkanoates such as acetate, propionate, tertiary butylacetate, cyclopentylpropionate, dimethylacetate, trirnethylacetate and phenoxyacetate; aryl esters such as benzoate, thiophene carboxylate, nicotinate; esters from dibasic organic esters such as the succinate, phthalate, and the sulfobenzoate; and those from polybasic inorganic acids such as sulfate and phosphate.

Our novel compounds are prepared by subjecting a l6-1ower alkyl-l,4-pregnadiene possessing a halogen substituent greater than fluorine at 0-6 to the action of zinc in alcohol or magnesium in ether or tetrahydrofuran. By proper control of acidity and solvent polarity, the 1,5-pregnadiene compounds are formed and isolated. Alternatively, the substituent at C6 may be a pseudohalogen such as tosylate or mesylate or a lower alkanoyloxy group like acetoXy or propionoxy. When the reactive group in the 6-position is halogen or tosylate, the reaction towards formation of the 1,5-dienes may proceed smoothly at room temperature. With a mesylate or lower alkanoyl ester in the 6-position, higher temperatures in the range of 50-l00 C. are preferred to eilect the transformation since at lower temperatures the reaction proceeds quite slowly.

The starting materials, that is, the 16-lower alkyl-l,4- pregnadienes are preferably prepared as described in copending application Serial No. 733,843 of Rausser et al., filed May 8, 1958. Introduction of the halogeno, pseudohalogeno or acyloxy substituent at C 6 is carried out by any one of several methods. For example, allylic halogenation of a 3-keto-l6-alkyl-1,4-pregnadiene with a reagent such as N-bromosuccinimide, N-bromoacetamide, N-chlorosuccinimide, bromine or chlorine in such solvents as dimethylsulfoxide or dioxane yields the corresponding 6-halogeno-16-alkyl-1,4-pregnadiene. Esterification of a 6-hydroXy-16-alkyl-1,4-pregnadiene can be made to yield the tosylate or mesylate or lower alkanoate intermediates. These 6-hydroxy-l6-alkyl-1,4- pregnadiene starting compounds are prepared by subjecting a o-desoxy-l6-alkyl-1,4-pregnadiene to the microbiological oxygenating action of an organism of the genus Chaetomium according to the analogous procedure described in Belgian Patent No. 548,450.

Prior to introducing a substituent to the '6-position of a l6-alkyl-1,4-diene so as to prepare the immediate precursor of the compounds of our invention, we prefer to esterify any reactive hydroxyl groups which may be present, such as C-ll or C-21. Thus, for example, in preparing 6-bromo-l6a-methylpredn=isone for ultimate conversion into l6a-methyl-l,5-pregnadiene-l7a,2l-diol-3,l1, ZO-trione, we prefer to employ a 21-ester o-f l6a-Inethylprednisone, such as the acetate, as the starting substance. In those instances where there is a hydroxyl group at C-ll such as when a loot-methylprednisolone analog is employed as a starting substance or an intermediate, the conditions required for esterification of the C-11 group of necessity esterify both the 17erand 21-hydroxy groups as Well. Thus when preparing a 6-halogeno-l6a-methylprednisolone derivative for use as an immediate precursor of a compound of this invention, an 11,17,21-triester such as l6u-methylprednisolone-ll,l7,2l acetate is employed. It is only in the particular instance where a halogenation step follows that esterification of a Cl1 hydroxyl group is advisable. However, if the intermediate being prepared is an ester such as 6-alkanoate or a 6-mesylate or a 6-tosylate, then it is only necessary to protect the hydroxyl group at C2l, since the esterification at C-6 can be carried out selectively in the presence of both an 11- and l7-hydroxyl group.

It is apparent that the l 6-alkyl-l,5-pregnadiene ob tained will of necessity contain the original ester protective groupings. These ester groups may be conveniently hydrolyzed either chemically by the use of dilute acid or microbiologically by means of Flavobacterium dehydro genans according to analogous procedures described in South African Patent No. 3,462/55. The polyhydroxy- 1,5-pregnadiene thus obtained may be selectively esterified at C-21 by methods well known in the art.

Alternatively, saponification of the protective ester groups may be carried out immediately after introducr tion of the substituent at C-6 and prior to conversion to a 1,5-diene. This process is applicable only to those methods wherein the intermediary compound contains a halogen at C6. The saponification of the ester may be efiected by means of a strong acid such as sulfonic, perchloric, or p-toluenesulfonic in aqueous alcohol. By way of example, 16a-methylprednisone is esterified to yield =1=6oc-methylprednis0ne 2l-acetate, whereupon bromine is introduced at C6. The 6-bromo-l6a-methyl prednisone 2l-acetate may be saponified to form 6-brorno- 16oc-methylprednis0ne which is then converted into the A -analog by reacting said 6-brom0-l6a-methylprednisone with zinc in alcohol yielding l6a-methyl-1,5-pregnadiene-l 711,2 l-diol-3,l l-20-trione.

Our process is applicable in general to the conversion of 16-lower alkyl-l,4-pregnadienes to l'6-lower alkyl-l,5- pregnadienes provided the starting 16-alkyl steroid c0ntains also a 3-keto-A moiety. Thus the preparation of 3-keto-9u,l 1 fl-dihalogenol6-alkyll ,S-pregnadienes from the corresponding 3-keto-9a,llfi-dihalogeno-ldalkyl-l,4 pregnadienes may be effected. Specifically, 9a,11fi-dichloro-l6cz-methyl-ll-desoxyprednisone 2l-acetate (prepared according to co-pending application, Serial No. 743,492, now Patent No. 2,894,963, filed June 20, 1958) is brominated to the requisite intermediate, 6-bromo- .Qmllfi dichloro 16:: methyl l1 desoxyprednisone ZI-aeetate, which, when reacted with zinc in alcohol, is convertible to 904-11B dichloro 16oz methyl 1,5 pregnadiene-17a,21-diol-3,2'0-dione ZI-acetate.

The 9,11 dihalogeno 1-6 alkyl 1,5 pregnadienes thus prepaIed by our process are potent anti-inflammatories, being particularly useful in topical administration.

Our novel -16-alkylated-1,S-pregnadienes are potent anti-inflammatory agents which are devoid of sodium or water retaining properties even when administered in large doses. It has been found that shifting the A -bond of prednisone or prednisolone gives rise to agents which exhibit a decrease in nitrogen, calcium and phosphorous loss, sodium and water retention, and. exhibit less increase in fasting blood sugar than occurs with administration of the 1,4-diene. Thus, although the side effects of prednisone and prednisolone are diminished by shifting the X -bond to C-5, these effects are still present. We have discovered that the introduction of the l6-alkyl group into a 1,5-diene completely eliminates the residual salt and water retaining properties of the substance, while simultaneously retaining the other advantageous properties, i.e. lower nitrogen, calcium and phosphorous loss and less increase in fasting blood sugar. Thus, since our novel compounds exhibit no water or sodium retention, they may be taken in large doses if necessary, providing for anti-inflammatory action without being concerned with edema-tons effect.

Our therapeutically valuable compounds are preferably administered orally in the form of tablets containing, for example about 2 to mg. per tablet mixed with a solid carrier containing one or more of the usual excipients such as starch, sugar, gums, soaps, clays and the like. Where parenteral administration is indicated, subcutaneous or intramuscular injection of a 2l-lower alkanoyl ester dissolved or suspended in a suitable nontoxic liquid vehicle is preferred. In the treatment of skin conditions such as atopic dermatoses, topical preparations such as containing 25% of active ingredient are advantageously employed. In some instances, such as direct treatment of an inflamed joint, crystalline suspensions are injected intra-articularly. These microcrystalline suspensions are adaptable for use in nasal sprays as well.

All the compounds of our invention are valuable therapeutic agents as outlined above. The preferred embodiment of our invention, however, are the 16mmethyl and 16l9-methyl-1,5-pregnadienes falling within Formula I, particularly the 9a-fluoro analogs of these 16-methyl-1,S- regnadienes.

The present invention is a continuation-in-part of our co-pending application Serial No. 725,521 filed April 1, 1958, now abandoned.

The following examples are illustrative of the procedure employed in preparing the compounds of this example, but are not to be construed as limiting the scope thereof; the scope of our invention being limited only by the appended claims.

EXAMPLE 1 9wflu0r0-16a-methyl-1 15,1 7a,21-trihydr0xy-1,5- pregnadiene-3,20-di0ne 21 -acetate A. GB-BROMO-Da-FLUORO-l6a-METHYL-116,17a,21-TRIHY- DROXY-l,4-PREGNADIENE-3,20-DIONE ZLACETATE The requisite starting material, 90t-flUOI'O-l6tx-ll16llhl 1lB,17a,2l-trihydroxy 1,4 pregnadiene-3,20-dione 21- acetate, is prepared by the process described in 00- pending application Serial No. 733,843 of Rausser et al filed May 8, 1958.

9a-fluoro t6u-methyl 11B,17u,21 trihydroxy-l,4-pregnadiene-3,20-dione. 21-acetate (380 mg.) is dissolved in 45 ml. of dioxane and then 165 mg. of bromine in ml. of dioxane is added to the stirred solution at room temperature. After the color is completely discharged, agitation is continued for another ten minutes before pouring the solution into 500 ml. of ice-water. The precipitate which thus forms is filtered and air dried, and is substantially 6 8-brorno-9e-fluoro-16a-methyl-11,9,

17a,21-trihydroxy-l,4-pregnadiene-3,20-dione 21- acetate.

This product is used without further purification in the following procedure.

B. 9a-FLUORO-ltFa-METHYL-llfifl7a.,21-TRIHYDROXY-L5- PREGNADIENE-3,20-DIONE ZI-ACETATE 65-bromo-9a-fluorod6a-methyl-11 8,17a,Z1-trihydroxy- 1,4-pregnadiene-3,20-dione 2l-acetate, prepared in above Example 1A, is dissolved in ml. of ethanol to which 15 ml. of water is added and the resulting solution is heated to reflux temperature. Zinc powder (1.5 g.) is then added and the refluxing suspension is stirred for 1.5 hours, cooled and filtered. The filtrate is concentrated to dryness in 'vacuo, and chromatographed over 30 g. of Floris-i1. The fractions eluted with benzene-ether (3:1) give 9a,-fiuoro-l6u-methyld1fi,17a,21-trihydroxy- 1,5-pregnadiene-3,ZO-dione 21-acetate.

EXAMPLE 2 9a-flu0r0-16ct-methyl-1 1 13,1 70,21 -trihydroxy-1 ,5 pregnadiene-3,20-di0ne The ldu-methyl-1,5-pregnadiene 2l-acetate prepared in above Example 1 is hydrolyzed to the corresponding 21- hydroxy compound with the aid of a culture of Flavobacterium a ehydrogenans (Rutgers University Collection No.

The culture of the organism is prepared by propagating it in a nutrient agar medium at 30 C. for 24 to 72 hours. During incubation, the inoculated tube is exposed to light with the resultant development of a yellow pigment characteristic of the species. The developed culture is rinsed from an agar slant under sterile conditions into a sterile medium of pH 6.8 and having the following composition:

Grams Yeast extract (Difco) 10 Potassium phosphate monobasic 4.48 Sodium phosphate dibasic 4.68

Tap Water to 1 liter.

This culture medium has previously been autoclaved, at 15 lb. pressure, for twenty minutes to obtain aseptic conditions, and cooled. The variant is grown in the medium under constant illumination, using the visible range of the spectrum. The incubation temperature is maintained at about 33 C. and is conducted under aerobic conditions. Aeration is accomplished by agitation and/ or blowing air through the culture medium.

After the organism has grown for 12 to 24 hours (or longer, if desired), 100 ml. of the growing culture are introduced into each of ten flasks, and to each flask are added 200 mg. of 9u-fluoro-16a-methyl-11fi,17a,21-trihydroxy-l,5-pregnadiene-3,20-dione 21-acetate dissolved in a minimum volume of ethanol. The reaction mixtures are then shaken at 30 C. for 12 to 72 hours. The reaction is stopped when paper chromatography indicates that there is no more starting material.

The contents of the flasks are combined and extracted with methylene chloride. The extracts are concentrated and the residue is crystallized from acetone-hexane yielding 9a-fluoro-16a-methyl-l 1 3,17a,21-trihydroxy-1,5-pregnadiene3,20-dione.

EXAMPLE 3 The requisite intermediate, 9a-fluoro16)8-methyl-1lfl, 17 a,2l trihydroxy- 1 ,4pregnadiene-3,20-dione 21-acetate, is prepared bythe process described in co-pending application Serial No. 733,843 of Rausser et al., filed May 8,

9a-fluoro-16p-methyl 11B,17oc,2l trihydroxy-1,4-pregnadiene-b,20-dione ZI-acetate (380 mg.) is brominated and the resulting product isolated in the manner described in Example 1A to give 6fi-bromo-9a-fiuoro-16B-methyl- 11B,17:1,2l-trihydroxy-l,4-pregnadiene-3,2O-dione 21-acetate.

B. 9a FLUOROl GB-METHYL-l118,17a,21-TRIHYDROXY-l,5-

PREGNADIENE-3.20-DIONE 2l-ACETATE EXAMPLE 4 The 16B-methyl-1,5-pregnadiene 21-acetate of above Example 3B is converted to the corresponding 2l-alcohol with the aid of a culture of Flavobacterium dehydrogenans in the manner described in Example 2 yielding 9u-fluoro-l6B-methyl 11fi,17oc,21 trihydroxy-1,5-pregnadiene-3,20-dione.

Alternatively, the 16,8-methyl-11 3,17a,21-trihydroxy- 1,5-pregnadiene of this example may be prepared as follows:

507 mg. of 6B-bromo-9a-fluoro-16a-methyl-11 8,171,21- trihydroxy 1,4 pregnadiene-3,20-dione 21-acetate (prepared as in Example 1A) is dissolved in 25 ml. of icecold ethanol containing 2.5 ml. of water and 65 mg. of sodium hydroxide. The solution is stirred under nitrogen at C. for fifteen minutes, then neutralized with acetic acid and concentrated in vacuo almost to dryness to yield a residue of substantially 65-bromo-9a-fluoro- 16,8-methyl-llfl,l7e,2l-trihydroxy 1,4-pregnadiene-3,20- dione. To this 65-bromo residue there is added icewater and ethanol and the solution is reacted with zinc in the manner of Example 1B to give 9a-flIlOI'O-l6flmethyl-l1/3,17a,21-trihydroxy 1,5 pregnadiene-3,20-dione.

EXAMPLE 9a-clzl0r0-16a-methyl-1JB,17u,21-trihydr0xy-1,5- pregnadiene-3 ,2 O-dione A. Gfi-BROMO-Qa-CHLORO-IGchMETHYLdl/El,170.,21-TRIHY- DROXY-l,4-PREGNADIENE-320-DIONE ZLACETATE The requisite intermediate, 9a-chloro-l6u-methyl-l 1 8, 171x,2l-trihydroxy-1,4-pregnadiene-3,20-dione ZI-acetate, is prepared in the manner described in co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

ot-ChlOI'OJGoc-IllfithYl-l1/3,17a,2l-trii1YdI'OXY-l,4 pregnadieue-3,20-dione 2l-acetate is bromina-ted, and the resulting product isolated in the manner of Example 1A to give 6,8-bromo-9a-chloro-16a-methyl-1lfi,17a,2l tr-ihydroxy-1,4-pregnadiene,3,20-dione Zl-acetate.

B. Qa-CHLORO-l6e-METHYL-l113,17a,2l-TRIHYDROXY1,5- PREGNADIENE-3,20-DIOI-IE ZI-ACETATE The 6fl-bromo-pregnadieneof above Example 5A is reacted with zinc and aqueous ethanol in the manner of Example 1B, and the resulting product isolated to give 90:- chloro Mot-methyl 1lfi,17 x,21 trihydroxy-1,S-pregnadiene-3,20-dione ZI-acetate.

c. 9a CHLORO-16a-METHYL-11B,l7a,21-TRIHYDROXY-1,5- PREGNADIENE-3,20DIONE The ZI-acetate of the 9a-chloropregnadiene prepared in above Example 5B is converted to the corresponding Zl-hydroxy with the aid of a culture of Flavobacterium' dehydrogenans to yield 9OL-Ch1OI'O-160tethyl-11 6,17a,21- trihydroxy-l,5-pregnadiene-3,20-dione.

Alternatively, the compound of this example may be prepared from 6 8-bromo-9a-chloro-16a-methyl-l15,170, 21-trihydroxy-1,4-pregnadiene-3,20-dione 2l-acetate (pre pared as in Example 5A) in the manner described in the alternative procedure of Example 4 whereby the ZI-acetate of the 6/3-bromo-1,4-pregnadiene is converted to the corresponding 21-hydroxy which, in turn, is reacted with zinc in ethanol-water to give 9a-ChlO-I0-l6oL-II16lZhYl-11fi,l7ot,2ltrihyd'roxy-l,5-pregnadiene-3,ZO-dione.

EXAMPLE 6 9a-chl0r0-16fi-methyl-11 3,1 7a,.21 -trihydr0xy-1 ,5- pregnadiene-3,20-di0ne A. 6B-BROMO-9 a-CHLORO-l GB-METHYL-l1B,17a,21-TRIHY- DROXY-1,4-PREGNADIENE-3,20-DIONE 21-ACETATE The requisite intermediate, 9a-chloro-16fi-methyl-11,3, 17oz,2l-trihydroxy-1,4-pregnadiene-3,ZO-dione 2 l-acetate is prepared according to the procedure described .in co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

The 9oc-Chl0r0 16,8-methyl-llfi,17a,21-trihydroxy-l,4 pregnadiene-3,20-dione 21-acetate is brominated and the resulting product isolated in the manner of Example 1A to give 6fl-brorno-9a-chloro-16,8-methyl-11fl,17o,21-trihy droxy-l,4 pregnadiene-3,20-dione ZI-acetate.

The 6B-bromo-9a-chloro-1,4-pregnadieneof above Example 6A is reacted with zinc powder in ethanol-Water in the manner of Example 1B to give 9a-chl0r0-16fi-methyl- 1 1B, 17:1,2 l-trihydroxy-1,S-pregnadiene-3,20-dione 21-acetate.

C. 9a-CHLORO-1 Sfl-METHYL-l113,17a,21-TRIHYDROXY-l,5- PRE GNADIEN'E3,20-DIONE EXAMPLE 7 9a-flu0r0-16a-metlzyl-17u,21-dihydr0xy-1,5- pregnadiene-3J1 ,ZO-trione A. 63 BROMO 9a FLUORO 16a METHYL 17a,21 DI- I-IYDROXY-I,4PREGN ADIEN E-3,l1,20-TRION E 2lACE- TATE The requisite intermediate, 9afiuoro-l6aamethyl-l7a, ZI-dihydroxy-1,4-pregnad'iene-3,11,20-trione ZI-acetate is prepared in the manner described in co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

9u-fiuoro-1 6a-methyl-17a,21-dihydroxy 1,4 pregnadiene-3,l1,2(l-trione 21-acetate is brominated, and the re sulting product isolated in the manner described in Example lA to yield 6 8-'bromo-9a-fiuoro-16u-methyl-l7u,21- dihydroxy- 1 ,4-pregnadiene-3,1 1,20-trione 21-acetate.

PREGNADIENE-3,11,20-TRIONE 21-ACETATE The 6,!3-bromo-1,4 pregnadieneof above Example 7A is reacted with zinc powder in aqueous ethanol in the manner of Example 13 and isolated, and purified in the described manner to give 9a-fluoro-16u-methyl-l7a,21-dihydroxy-1,5-preguadiene-3,11,20-trione ZI-acetate.

PREGNADIENE-3,11,20-TRIONE The 9wfluoro-1,S-pregnadiene-Zl-acetate of Example 73 is converted to the corresponding '21-hydroxy by the action of a culture of Flavobacterium dehydrogenans in the manner of Example 2 to yield 9ot-fluoro-16u-methyl- 1705,21-(lil13'd1'0XY- 1,5-pregnadiene-3 ,1 1,20-trione.

Alternatively, the compound of this example may be prepared in the manner described in the alternative procedure of Example 4, whereby 6,8-brm0-9oc-fiuOrO-l6ocmethyl-17mll-dihydroxy-l,4-pregnadiene-3,11,20 trione 21-acetate (compound of Example 7A) is first hydrolyzed to the corresponding 21-alcohol, then reacted with zinc to yield 9a-fluoro-16a-methyl-17u,2l-dihydroxy-l,S-pregnadiene3,1l,20trione.

EXAMPLE 8 9e-flu r0-l6B-methyl-17a,21-dihydr0xy,1,5- pregnadiene-3 ,1 1,20-tri0ne A. 65 BRONEO 9a FLUORO 16B METHYLl7a,2l-DI- HYDROXY 1,4 PREGNADIENE 3,11,20 TRIONE Zl-ACETATE The requisite intermediate, 9a-fiuoro-16 3methyl-17a, 2l-dihydroxy-1,4-pregnadiene-3,11,20-trione 21-aceta-te, is prepared by the process described in co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

In the manner of Example 1A, 9a-fiuoro-16fi-methyl- 17a,21-dihydroxy-1,4-pregnadiene-3,1 1,2D-trione 21-acetate is brominated and the resulting product isolated to give 6fl-bromo-9a-fluoro-16fl-methyl-17a-21-dihydroxy-1, 4-pregnadiene-3 ,1 1 ,ZO-tn'one 21-acetate.

PREGNADIENE-3,11,20-TRIONE fill-ACETATE In the manner of Example 1B, the 6,13-bro-mo-1,4-preg nadiene of above Example 8A is reacted with zinc in aqueous ethanol to give 9oc-fiuor0-16B-methyl-17a,21-dihydroxy-l,5-pregnadiene-3,11,20-trione 21-acetate.

PREGNADIENE-3,11,204DRIONE In the manner of Example 2, the 9oc-iflu0r016,B-methyl- 1,5-pregnadiene 2l-acetate of Example 813, is converted to the corresponding 21-alcohol by the action of a culture of Flavobactcrium dehydrogenans to give 9a-fluOIO-16B- methyl-17a,21-dihydroxy-1,5-pregnadiene-3 ,11,20-trione.

Alternatively, the compound of this example may be prepared by the alternate procedure of Example 4 from 6fl-bromo-9a-fiuoro-16,B-methyl-17a,21 dihydroxy 1,5- pregnadiene-3,l1,20trione 21-acetate (the compound of Example 8A) to give 9a-flu0rol6B-methyl-17a,2l-dihydroxy-1,5-pregnadiene-3 ,11,20-trione.

EXAMPLE 9 Qa-bromO-I 6 8-methyl-1 118,1 7a,21-trihydroxy,1,5- pregnadiene-3,Z0-di0ne A. 613,90. DIBROMO 16B METHYL 11B,17a,21 TRI- I-IYDROXY 1,4- PREGNADIENE 3,20 DIONE 2l- ACETATE In the manner of Example 13, the 6;3,9 :t-dibromo-1,4-v

pregnadiene of Example 9A, is reacted with zinc in aqueous ethanol and the resulting product isolated and purified in the described manner to give 9a-br0mo-16B- methyl-115,17u,21-trihydroxy-1,5-pregnadiene-3,2O dione ZI-acetate.

l C; 9a-BROMO-IGB-METHYL-llfi,17u,21-TRIHYDROXY-1,5- PREGNADIENE-3,20-DIONE The 21-acetate of Example 913 is converted to the corresponding 2l-alcohol With the aid of a culture of Flavobacterium dehydrogenans in the manner of Example 2 to give 9oz-bromo-16fl-methyl-11B,17a,21-trihydroxy-1,5- pregnadiene-Z,20-dione.

Alternatively, the compound of this example may be prepared by the alternative procedure of Example 4 from 6,8,9a-dibromo 16 3 methyl 11fi,17a,2l-trihydroxy-1,4- pregnadiene-3,20-dione 21-acetate to give 9a-bromo-16fimethyl-1 1p],17a,21-trihydroxy-1,5-pregnadiene3 ,ZO-dione.

EXAMPLE 10 9a bromo 16a methyl 11p,17u,21 trihydroxy 1,5-

pregnadiene-3,20-di0ne A. 6B,9a-DIBROMO-1Ga-lVIE'lHYL-llfi,17a,2l-TRIHYDROXY- 1,5-PREGNADIENE-3,20-DIONE Zl-ACE'IATE The requisite intermediate, 9a-bromo-l6a-methyl-11fi, 17:1,21-trihydroxy-1,4-pregnadiene-3,20-dione 21-acetate is prepared by the process of copending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

The above bromo intermediate is brominated, and the resulting product isolated in the manner of Example 1A to yield 66,9oz-dibromo-16u-rnethyl-115,l7a,21-trihydroxy- 1,4-pregnadiene-3,20-dione 21-acetate.

B. 9a-BROMO-lGa-METHYLd113,17a,21 TRIHYDROXY-1,5-

PREGNADIENEQflO-DIONE 2l-ACETATE In the manner of Example 1B, the 6B,9oc-dibrorno1,4- pregnadiene of Example 10A is reacted with zinc in aqueous ethanol to give 9u-bromo-16m-methyl-11B,17m,21- trihydroxy-1,5-pregnadiene-3 ,20-dione 21-acetate.

C. Qa-BROMO-lfia-METHYL 11B,17,21-TRIHYDROXYJ.,5- PREGNADIENE3,20DIONE The 21-acetate pregnadiene of Example 10B is hydrolized to the corresponding 21-alcohol with the aid of a culture of Flavobacterium dehydrogenans in the manner of Example 2 to give 9a-bromo-16a-methyl-11 8,17a,21 trihydroxy-1,5-pregnadiene-3,20-dione.

The compound of this example may also be prepared from 618,9a-dibr0mo-16a-methyl-l 15, 17a,21-trihydroxy-1 4-pregnadiene-3,ZO-dione 21-acetate (the compound of Example 10A by the alternate process of Example 4 yielding 9a-bromo-16a-methyl-llfl,l7a,21-trihydroxy-1,5- pregnadiene-3,20-dione.

EXAMPLE 11 9a broma 16oz methyl 17a,21 dihydroxy 1,5 pregnudism-3,11 ,ZO-trione A. 6B,9a-DIBROMO-l6wMETHYL-l7a,21-DIHYDROXY1,-1- PREGNADIENE-3,11,20'TRIONE 21-ACETATE The requisite intermediate, 9a-br0m0-16a-methyl-17a,

I ZI-dihydroxy-l,4-pregnadiene-3,l1,20-trione 21-acetate, is

prepared according to the procedure in co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

This intermediate is brominated in the manner of Example 1A and the resulting product isolated in the described manner-to give 65,9a-dibromo-16ot-methyl-17a,21- dihydroxy-1,4-pregnadiene-3,11,20-tri0ne 2l-acetate.

B. 9wBROMOJ.6a-METHYL-l7a,21-DIHYDROXY-1,5- PREGNADIENE-3,l1,ZO-TRIONE ZI-ACETATE The dibromide of Example 11A is reacted with zinc in aqueous ethanol in the manner of Example 1B, and the resulting product isolated and purified in the described manner to give 9a-bromo-lfia-rnethyl-17a,21-dihydroxy- 1,5-pregnadiene-3,l1,20-trione 21-acetate.

C. 9a-BROMO-l6a-METHYL-l7a,21-DIHYDROXY-1,5- PREGNARIENE-3,11,20-TRIONE In the manner described in Example 2, the ZI-acetate of above Example 11B is hydrolyzed to the correspond- EXAMPLE 12 9a bromo 16,8 methyl 17o,21 dihydroxy 1,5- pregnadiene-3,ILZO-trione A. 65,9a-DIBROMo-1(in-METHYL-l7a,21-DIHYDROXY-1,4- PREGNADIENE-3,11,20-TRIONE 2l-ACETATE The requisite intermediate, 9ot-br0mo-1618-methyl-l7e, 2l-dihydroxy-1,4-pregnadiene-3,l1,20-trione ZI-acetate is prepared by the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

The 9a-bromopregnadiene intermediate is brominated and isolated in the manner of Example 1A to give 6 8,90- dibromo-16fi-methyl-17a,21 dihyclroxy 1,4-pregnadiene- 3,11,20-trione ZI-acetate.

B. Qa-BR OMO-l GB-METHYL-17a,2lDIHYDROXYl,5 PREGNADIENE-3,l1,20-TRIONE 21-ACETATE The dibromide of Example 12a is reacted with zinc powder in aqueous ethanol in the manner of Example 1B, and the resulting product isolated and purified in the described manner to give 9a-bromo-16fi-methyl-17a,2l-dihydroxy-1,5-pregnadiene-3,11,20-trione 2 l-acetate.

C. Qa-BROMO-l(SB-METHYL-17:1,21-DIHYDROXY-L5- PREGNADIENE-3,11,20-TRIONE The ZI-acetate of Example 128 is hydrolyzed to the corresponding alcohol with the aid of a culture of Flavobacrerilmz dehydrogenans to yield 9wbromo-16/3-methyl- 170:,21-dihydroxy-1,5-pregnadiene-3,11,20-trione.

EXAMPLE 13 160: methyl 11p,17a,21 trihydroxy 1,5 pregnadiene- 3,20-dine G/SBROMOJ6aMETHYL-11B,170.,2LTRIHYDROXY-L4- PREGNADIENE-3,20-'DIONE 2l-ACETATE The requisite intermediate, 16a-methyl11B,17a,21-trihydroxy-l,4-pregnadiene-3,ZO-dione ZI-acetate, is prepared by the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

The 16a-methyl-l,4-pregnadiene intermediate is brominated and the resulting product isolated in the manner of Example 1A to give 6fi-bromo-16a-methyl-l1/8,17a,21-trihydroxy-1,4-pregnadiene-3,20-dione Zl-acetate.

DIENE8,20-DIONE 21-ACETATE The 6/3-bromo-L4-pregnadiene of Example 13A (500 mg.) is dissolved in 159 ml. of absolute ethanol, then 25 ml. of Water and g. of zinc powder are added. The suspension is stirred at room temperature for 8 hours, after which time the zinc is filtered. The filtrate is concentrated in vacuo to a residue which is crystallized from acetone to give l6a-methyl-11B,17u,21-trihydroxy-1,5- pregnadiene-3,20-dione 2l-acetate.

Alternatively, the compound of this example is prepared from the 6fi-brorno-1,4-pregnadiene of Example 13A with zinc in ethanol by the procedure of Example 1B.

C. lBa-METHYL-llfi,17a,21-TRIHYDROXY-l,5 PREGNADIENE-3,20-DIONE The Ill-acetate of above Example 13B is hydrolyzed to the corresponding 21-alcohol with the aid of a culture of Flavobacteriztm dehydrogenans in the'manner of Example 2 to give 16a-methyl-115,17a,21-trihydroxy-1,5-pregnadiene-3,20-dione.

Alternatively, the product of this example may be preit pared from 6fl-bromo-l6a-methyl-11fl,17a,21-trihydroxyl,4-pregnadiene-3,ZO-dione ZI-acetate by the alternative process of Example 4.

EXAMPLE l4 16/8-methyl-J 1,8,1 7a,21-trihydroxy-1 ,5 -pregnadiene- 3,20-aione A. 6B-BROMO 165-METHYL-1 16,17a,2'1-TRIHYDROXY1,4- PREGNADIENE-3,20-DIONE 21-ACETAIE The requisite intermediate, 16fl-methyl-1lB,17u,21-trihydroxy-1,4-pregnadiene-3,ZO-dione ZI-acetate, is prepared according to the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

This 1Sfi-methyl-1,4-pregnadiene intermediate is bromin-ated, and the resulting product isolated and purified in the manner of Example 1A to give 6B-bromo-16fimethyl 11,8,17oz,21 trihydroxy 1,4 pregnadiene 3,20- dione 21-acetate.

The fi-bromo-lA-pregnadiene of Example 14A is reacted With zinc powder in aqueous ethanol in the manner of Example 13B, and the resulting product isolated and purified in the described manner to give 16fl-methyl-11B, 171,2l-trihydroxy-1,5-pregnadiene-3,20-dione 21-acetate.

The 21-acetate of Example 148 is hydrolyzed to the corresponding 21-alcohol with the aid of a culture of Flavobacterium dehydrogenans in the manner described in Example 2 to give 16,B-methyl-11/i,17a,21-trihydroxy- 1,5-pregnadiene-3,ZO-dione.

Alternatively, the compound of this example may be prepared from 6 8-bromo 16,8 methyl-ll,8,l7a,2l-trihydroxy-l,4-pregnadiene-3,ZO-dione '2l-acetate (the compound of Example 14A) by the alternative process of Example 4.

EXAMPLE 15 16u-methyl-1 70,21-dihydr0xy-1,S-pregnadiene- 3,11,20-trione A. 6flBROM0 16a-METHYL 1 7a,21-DIHYDROXY-1,4 PREGNADIENE-E,11,20-TRIONE Zl-ACETATE The requisite intermediate, 16a-methyl-17a,21-dihydroxy-l,4-pregnadiene-3,11,20-trione 21-acetate, is prepared according to the process of .co-pending application Serial No. 733,843 otRausser et al., filed May 8, 1958.

This 1Gm-methyl-1,4-pregnadiene intermediate is brominated and the resulting product isolated in the manner of Example 1A to give 6B-bromo-16a-methyl-17a,21-dihydroxy-l,4-pregnadiene-3,11,20-trione 21-acetate.

.B. 16a-METHYL-17a,2'1-DIHYDROXY-1,5PRJEGNADIENE- 3,11,20-TRIONE ZI-ACETATE The 6B-brorno-1,4-pregnadiene ofExample 15A is reacted with zinc powder in aqueous ethanol in the manner of Example 133, and the resulting product isolated and purified in the described manner to give l6a-methyl-17a, 2l-dihydroxy-1,5-pregnadiene-3,1 1,20-trione 21-acetate.

C. 1Ga-METHYL-l'ia,21-DIHYDROXY-l,fi-PREGNADIENE- 3,11,20TRIONE The 2l-acetate of Example 15B is hydrolyzed to the corresponding 21-alcohol with the aid or" a culture of F lavobacterium dehydrogenans in the manner described in Example 2 to give Mot-methyl-17u,21-dihydroxy-1,5- pregnadiene-3,l1,20-t1i0ne.

Alternatively, the compound of this example may be prepared from 6B- bromo-16a-methyl-l7cx,2l-dihydroxyl,4-pregnadiene3,11,20-trione ZI-acetate (the compound of Example 15A) by the alternative process of Exam ple 4. Y

1 1 EXAMPLE l6 16y3-methyl-1 7a,.21-dihydrwcy-1 ,5 -pregnadiene- 3,11 ,ZO-trione A. Gfl-BROMO-l6fll\IETHYL-17n,21-DIHYDROXY-1,4- PREGNADIENE-3,11,20TRIONE ZL-ACETATE The requisite intermediate, l6 3-methyl-17a,21-dihydroxy-1,4-pregnadiene-3,11,20-trione ZI-acetate is prepared according to the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

This l6B-methyl-1,4-pregnadiene intermediate is brominated and the resulting product isolated in the manner of Example 1A to give 6B-bromo-l6fl-methyl-17u,21-'dihydroxy-1,4-pregnadiene-3,l1,20-trione 2 l-acetate. B. 1GB-METHYL-17a.,21-DIHYDROXY-1,5-PREGNADIENE- 3,11,20-TRIONE 21-ACELATE The GB-bromo-lA-pregnadiene of Example 16A is reacted with a Zinc powder in aqueous ethanol in the manner of Example 13B, and the resulting product isolated and purified in the described manner to give 16fl-methyll7a,2l-dihydroxy-1,5-pregnadiene 3,11,20 trione 21- acetate.

C. 1GB-METHYL-l70.,2'1-DIHYDROXYJ,S-PREGNADIENE- 3,11,20-TRIONE The 21-acetate of Example 16B is hydrolyzed to the corresponding 21-alcohol with the aid of a culture of q Flavobacterium dehydrogenans in the manner described in Example 2 to give 166-methyl-l7a,21-dihydroxy-1,5 pregnadiene-3,1 1,20-trione.

Alternatively, the compound of this example may be prepared from 6B-bromo-1fifi-methyl-l7u,21-dihydroxy- 1,4-pregnadiene-3,l1,20-trione Zl-acetate (the compound of Example 16A) by the alternative process of Example 4.

EXAMPLE 17 purified in the described manner to give 16a-6ihYl-l7ct,2ldihydroxy-l ,5-pregnadiene-3, l 1,20-trione 2 l-acetate.

C. 1(ia-ETHYL-l'?a,2l-DIHYDROXY-1,5PREGNADIENE- 3,11,20-TRIONE The ill-acetate of Example 178 is hydrolyzed to the corresponding 21-a1cohol with the aid of a culture of Flaw-bacterium dehydrogenans in the manner described in Example 2 to give 16a-ethyl-17c,21-dihydroxy-1,5-pregnadiene-3 ,1 1,20-trione.

Alternatively the compound of thi example may be prepared from 6fi-bromo-16a-ethyl-17u,2l dihydroxy-1,4- pregnadiene-3,11,20-trione 21-acetate (the compound of Example 17A) by the alternative process of Example 4.

EXAMPLE l8 16e-ethyI-11 [3,1 701,21 -trihydrxy-1 ,5 -pregnadiene- 3 ,2 O-dione A. GB-BROMO-l6a-ETHYL-11B,17 a,21-TRIHY'DROXY-1,4- PREGNADIENEEEWDIONE 21-ACET'ATE The requisite intermediate, l6a-ethyl-llfl,l7ot,2l-trihy droxy-l,4-pregnadiene-3,ZO-dione 2l-acetate, is prepared according to the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

This la-ethyl-l,4-pregnadiene intermediate is brominated and the resulting product isolated in the manner of Example 1A to give 6fl-bromo-16ot-ethyl-l1,8,17ot,21- trihydroxy-l,4-pregnadiene-3,ZO-dione 2l-acetate.

B. 16a,ETHYL 11B, 17a,2l-TRIHYDROXY-l,5-PREGNADI- ENE-3,20-DIONE 21-ACETATE The 6l3-bromo-L4-pregnadiene of Example 18A is reacted With a zinc powder in aqueous ethanol in the manner of Example 1B, and the resulting product isolated and purified in the described manner to give 16a-ethyl- 11B,l7e,2i-trihydroxy-1,5-pregnadiene-3 ,20-dione 21-acetate.

C. 16a-ETHYL-115,170.,2l-TRIHYDROXY-LS- PREGNADIENE-3,20-DIONE The 21-acetate of Example 188 is hydrolyzed to the corresponding Zl-alcohol with the aid of a culture of F lavobacterium dehydrogenans in the manner described in Example 2 to give l6a-ethyl-1lB,17a,21-trihydroxy- 1,5-pregnadiene-3,ZO-dione.

Alternatively, the compound of this example may be prepared from 6fi-bromo-16a-ethyl-l l B,17u,21-trihydroxy- 1,4-pregnadiene-3,ZO-dione 2l-acetate (the compound of Example 18A) by the alternative process of Example 4.

EXAMPLE l9 9a-bromo-16a-ethyl-11,B,1 7a,21lrihydr0xy-I,5-pregnadiene-3,20-dione A. 66,9a-DIBROMO-lGa-ETHYL,11B,l7a,21-TRIHYDROXY- 1, i-PREGNADIENE-3,20-DIUNE 21-ACETATE The requisite intermediate, 9or-bromo-16a-ethyl-1lp, l7a,21-trihydroxy-l,4-pregnadiene-3,20-dione Zl-acetate, is prepared according to the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

This 9a-bromo-l6a-ethyl-l,4-pregnadieneintermediate, is brominated and the resulting product isolated in the manner of Example 1A to give 618,9a-dibromo-l6a-ethyl- 11B,17a,2l-trihydroxy-l,4-pregnadiene-3,20-dione 2l-acetate.

B. (la-BROMO-lGa-ETHYL-l1/3,17a,21TRIHYDROXY1,5-

PREGNADIENEEQO DIONE 2-1-ACETATE The 6fl-bromo-L4-pregnadiene of Example 19A is reacted with a zinc powder in aqueous ethanol in the manner of Example 113, and the resulting product isolated and purified in the described manner to give 9a-bromo-16rxethyl-11}8,l7a,21-trihydroxy-1,5-pregnadiene 3,20 dione 21-acetate.

C. 9ct-BROMO-16a-ETHYL-115,1711.,21-TRIHYDROXY-L5- PREGNAD'IENE-3,20-DIONE The 2'1-acetate of Example 19B is hydrolyzed to the corresponding 21-a1cohol with the aid of a culture of Flavobacterium dehydrogenans in the manner described in Example 2 to give 9a-bromo-16a-ethyl-l1,B,17a,2l-trihydroxy-1,5-pregnadiene-3,ZO-dione.

Alternatively, the compound of this example may be prepared from 6 8,9a-dibromo-16wethyl-llfi,l7a,2l-trihydroxy-l,4-pregnadiene-3,ZO-dione 21-acetate (the compound of Example 19A) by the alternative process of Example 4.

EXAMPLE 20 9ot-flu0ro-16u-ethyl-11BJ7ot,21-trihydroxy-1,5-pregnadiene-3,20-dione A. Sfi-BROMO-Sa-FLUORO-lGa-EI'HYL-l15,170.,21-TR1HY- DROXY-1,4-PREGNADIENE-3,20-DIONE ZI-ACETATE The requisite intermediate, t-fiUOIO-16OL-6thyi-1 15,17rx, 2l-trihydroxy-1,4-pregnadiene-3,20-dione Zl-acetate, is prepared according to the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

This 9a-fiuoro-16a-ethyl-1,4-pregnadiene intermediate is brominated and the resulting product isolated in the manner of Example 1A to give 6 8-bromooc-flllOrO-16aethyl-1 l B,17a,2l-trihydroxy-1,4-pregnadiene 3,20 dione 21-acetate.

B. 9a-FLUORO-l6a-ETHYL-11fl,1Ta,21-TRIHYDROXY-1,5- PREGNADIENE-S,20-DIONE 2l-ACETATE The 6fi-bromo-9a-fiuoro-1,4-pregnadiene of Example 20A is reacted with zinc powder in aqueous ethanol in the manner of Example 1B, and the resulting product isolated and purified in the described manner to give 9a-fluoro-16ocethyl-115,17a,21-trihydroxy-1,5-pregnadiene 3,20 dione 21-acetate.

C. QmFLUORO-l6a-ETHYL11B,l7a,2l-TRIHYDROXY1,5- PREGNADIENE-3,20-DIONE The Zl-acetate of Example 20B is hydrolyzed to the corresponding 21-alcohol with the aid of a culture of Flavobacr'erizzm dehydrogenans in the manner described in Example 2 to give 9oz-filJ010-l6a-ethyl-l1,8,l7vz,2lt1lhydroxy-1,5-pregnadiene-3,ZO-dione.

Alternatively, the compound of this example may be prepared from 6B-bromo-9a-fluoro-16u-ethyl-l l/3,17a,21- trihydroxy-1,4-pregnadiene 3,20 dione 2l-acetate (the compound of Example 20A) by the alternative process of Example 4.

EXAMPLE 21 A. 66 BROBIO-9 a-FLUORO-1 6a-ETHYL-17a,21-DIHYDROXY- l,''l-PREGNADIENE-3,11,20TRIONE 2l-ACETATE The requisite intermediate, 9zx-fluO1016oc6thy1-170c,2ldihydroxy-1,4-pregnadiene-3,11,20-trione 2l-acetate, is prepared according to the process of co-pending application Serial No. 733,843, of Rausser et al., filed May 8, 1958.

This 9u-fiuoro-16a-ethyl-1,4-pregnadieneintermediate is brominated and the resulting product isolated in the manner of Example 1A to give 6,8-bromo-9a-fluorod6methyl-17a,21-dihydroxy-1,4-pregnadiene-3,11,20-trione 21- acetate.

B. 9a-FLUORO-16dETHYL-17a,2l-DIHYDROXY-l,5- PREGNADIENEB,11,20TRIONE 2l-ACETATE The 6fi-bromo-9a-fiuoro-1,4-pregnadiene of Example 21A is reacted with zinc powder in aqueous ethanol in the manner of Example 1B, and the resulting product isolated and purified in the described manner to give 90:- fluoro-l6oz-ethyl-l7a,21-dihydroxy-1,5-pregnadiene 3,11, ZO-trione 21-acetate.

C. 9a.FLUORO-16a-ETHYL-17a,21DIHYDROXY-l,5- PREGNADIENEBJlZO-TRIONE The 2l-acetate of Example 21B is hydrolyzed to the corresponding 2l-alcohol with the aid of a culture of Flavobacterium dehydrogenans in the manner described in Example 2 to give 9a-fluoro-16a-ethyl-l7a,21-dihydroxy-l,5-pregnadiene-3,l1,20-trione.

Alternatively, the compound of this example may be prepared from 6 3-bromo-9a-fluoro-16a-ethyl-17a,2l-dihydroxy-l,4-pregnadiene-3,11,20-trione 2l-acetate (the compound of Example 21A) by the alternative process of Example 4.

EXAMPLE 22 1ooz-n-butyl-l7a,21-dihydr0xy-I,5-pregnadiene-3,11,20- trione A. 6-BROMO1GwN-BUTYLJWZ,2lDIHYDROXY-1,4-

PREGNADIENE-3,11,20-TRIONE 2'1-ACETATE The requisite intermediate, 16a-n-butyl-l7a,21-dihydroxy-l,4pregnadiene-3,l1,20-trione 2l-acetate, is derived from 16rx-n-butylcortisone 21-acetate, which,'in turn, is

prepared according to the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958. l6a-n-butylcortisone 21-acetate is subjected to the action of a culture of Corynebacterium simplex (A.T.C.C. 6946) in the manner disclosed in US. Patent No. 2,837,- 464, and the resulting reaction mixture extracted with chloroform in the described manner. The chloroform extracts are combined and concentrated to a residue which, after crystallization from acetone, gives 16a-n-butyl-17a, 2l-dihydroxy-1,4-pregnadiene-3,11,20-trione 21-acetate.

16a-n-butyl-17 11,21dihydroxy-1,4-pregnadiene 3,11,20- trione 21-acetate is brominated and the resulting product isolated in the manner of Example 1A to give 6 9-bromol6a-n-butyl-l7u,21-dihydroxy-1,4-pregnadiene-3,11,20-trione 21-acetate.

B. 16a-N-BUTYL-17a,21-DIHYDROXY-1,5-PREGNADIENE 3,11,2O IRIONE ZLACETATE The 6,6-bromo-16a-n-butyl-1,4-pregnadiene of above Example 22A is reacted with zinc in aqueous ethanol in the manner of Example 1B and the resulting product isolated and purified in the described manner to give l6cx-I1- butyl17u,21-dihydroxy-1,5-pregnadiene-3,11,20-trione 21- acetate.

C. 1Ga-N-BUTYL-l7a,21-DIHYDROXY-l,S-PREGNADIENE- 3,11,20TRIONE The 2l-acetate of above Example 22B is hydrolyzed to corresponding 21-alcohol with the aid of a culture of Flavobacterium dehydlogenans by the procedure described in Example 2.

Alternatively, the compound of this example may be prepared from 6fi-bromo-l6u-n-butyl-l7a,21-dihydroxy-1, 4-pregnadiene-3,l1,20-trione Ill-acetate (the compound of Example 22B) by the alternative process of Example 4.

EXAMPLE 23 16,8-n-butyl-1 7a,21-dihydr0xy-1,5-pregnadiene-3,11,20- trione The requisite intermediate, 16fi-n-butyl-17a,21-dihydroxy-1,4-pregnadiene-3,11,20-trione ZI-acetate, is derived from 16B-n-butylcortisone 2l-acetate which is prepared by the process of co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958. When subjected to the action of a culture of Carynebacterium simplex in the manner disclosed in US. Patent No. 2,837,464, and the resulting product isolated and purified as described in EX- ample 22A, 16,6-n-butylcortisone 21-acetate is converted to 16fi-n-butyl-1711,21-dihydroxy1,4-pregnadiene-3,l1,20- trione 21-acetate.

In the manner of Example 22, 16;3-n-butyl-17a,21-di hydroxy-l,4-pregnadiene-3,11,20-trione 21-acetate is brominated to the corresponding 6fi-bromo-deriwative which, in turn, is reacted with zinc in aqueous ethanol to give 16,3-n-butyl l7oc,2l dihydroxy-1,5-pregnadiene-3,11,20- trione 2l-acetate.

The 2l-acetate is then hydrolyzed to the 21-alcohol in the described manner to give 16fi-n-butyl-l7a,2l-dihydroxy-1,5-pregnadiene-3,l1,20-trione.

EXAMPLE 24 16ot-methyl-17a,21-dihydr0xy-1,5-pregnadiene-3,11,20 trione 21 -n-pr0pionate To one gram of 16a-methyl-17a,21-dihydroxy-l,5-preg nadiene-3,ll,20-trione, the compound of Example 15, there is added 0.5 ml. of n-propionic anhyd-ride in 2.3 ml. of pyridine. After standing for one hour at room temperature, the mixture is poured into ice and hydrochloric acid. The resulting precipitate is filtered and recrystallized from aqueous methanol to yield 16a-methyl-17a,21- dihydroxy- 1 ,5-pregnadiene-3 ,1 1,20-trione 21-n-propionate.

Similarly, any one of the 2l-hydroxy-1,5-pregnadiene compounds of our invention may be treated with propionic anhydride in the above described manner to yield the corresponding 2l-n-propionate ester.

16m-mezhyl-1 70:,21-dihydr0xy-1,S-pregnadiene- 3,11 ,ZO-trione A. l6a-l1ETI-IYL 65,1711,2'1-TRIHYDRGXY-1A- PREGNADIENE-3,11,20-TRIONE The requisite intermediate, 16o: methyl 170;,21 dihydroxy-1,4-pregnadiene-3,11,20-trione, is prepared as described in copending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

Into each of ten 300 ml. Erlenmeyer flasks is placed 100 ml. of a growth medium having the following composition:

G. Difco yeast extract 3-10 Corn steep liquor 1 Dextrose l Distilled or tap Water, q.s. to one liter.

Each flask is inoculated with spores from an agar medium culture of Chaetomium funicolztm (QM No. 33C) or with a l-10% submerged inoculum which has been grown for 2448 hours. The mixture is incubated by shaking the flasks on a rotary shaker for 24-48 hours at 28 C. at approximately 250 rpm. To each of the flasks (now showing prolific growth) is added, aseptically, 50 mg. of 16a-methyl-l7a,2l-dihydroxy-1,4-pregnadiene- 3,11,20-trione in 2 ml. of ethanol. The fermentation mixture is then incubated and shaken for 24-72 hours at 28 C. after which time complete conversion occurs. The mixture is extracted thoroughly with chloroform and the small fraction of the product which is retained within the mycelium is extracted by boiling the mycelium in chloroform for a few minutes. The chloroform extracts are combined and evaporated to a residue yielding ap proximately 5 mg. of crude material. The residue is triturated with methanol affording a crystalline solid which is purified by crystallization from acetone yieldring 1611 methyl 6fl,l7ot,21 trihydroxy-l,4-pregnadiene- 3,1 1,20-trione.

B. 16a-l\IETHYL-6B,l7ct,21 TRIHYDROXY-1',4-PREG NADIENE-3,11,20-TRIONE 21-ACETATE 100 mg. of 16a-methyl-6fl,17a,21-trihydroxy-1,4-preg nadiene-3,1 1,20-trione 21-acetate, is prepared as in Example 25B, is dissolved in 0.4 ml. of methylene chloride and 0.4 ml. of pyridine. The solution is cooled to 0 C., 1.6 ml. of methanesulfonyl chloride added, and the mixture stirred for 2 hours at 0 C. then left overnight at room temperature. The reaction mixture is then poured into water, the resulting precipitate which is filtered and air dried is 16u-methyl-6e,17ot,21-trihydroxy-1,4-p-regnadiene- 3,11,20-tr-ione G-methanesulfonate 2l-acetate.

D. lfia-METHYLl'ia,21DIHYDROXY-1,S-PREGNADIENE- 3,11,20-TRIONE iii-ACETATE The 6 methanesulfonate 16a-methyl-lA-pregnadiene (55 mg.) of above Example 250 is dissolved in 20 ml. of

ethanol and 1 ml. of water. Zinc fillings (100 mg.) are added, and the suspension stirred for 1 hour at C. The solution is cooled, the zinc filtered, and the resulting filtrate concentrated to a residue of substantially 16amethyl-17a,21-dihydroxy 1,5-pregnadiene-3,11,20-trionc 21-acetate.

E. 16a-METHYL-l7a2l-DIHYDROXY-l,5-PBEGNADIENE =3,11,20-TRIONE The 21-acetate of Example 25D is hydrolyzed to the corresponding 2l-alcohol in the manner described in Example 2 to give 16u-methyl-17e,2l-dihydroxy-l,5-pregnadiene-3,11,20-trione.

Alternatively, the compound of this example may be prepared as described in following procedures F and G. F. 16a \IETHYL 65,170.,21 TRIHYDROXY 1,4 PREG- NADIENE-3,11,20-TRIONE 613,21-DIACETATE? A solution of 25 mg. of the compound of Example 25A in 2 ml. of anhydrous pyridine is poured onto mg. of acetic anhydride in an anhydrous atmosphere. The mixture is stirred for 30 minutes and then poured into dilute sulfuric acid and ice. The resultant precipitate is removed by filtration, dried and crystallized from methanol, yielding 16a methyl 6,6,17a,21 trihydroxy 1,4 pregnadiene-3,ll,20-trione 65,21-diacetate.

G. 16a-METHYL-17a,21-DIHYDROXY-l,5-PREGNADIENE- 3,11,20-TRIONE 2l-ACETATE The 6fl,21-diacetate of Example 25F is reacted with zinc and alcohol in the manner described in Example 25D to give 16a methyl 2l-dihydroxy-1,5-pregnadiene- 3,11,20-trione Zl-acetate.

The 21-acetate of this example is convented to the corresponding 21-hydroxy-l,S-pregnadiene in the manner of Example 2.

A third alternative process is described in following procedures H and I.

H. 1601. METHYL Gfi,17a,21 TRIHYDROXY 4,4 PREG- NADIIENE-3,l1,20-TRIONE GIS' p-TOLUENESULFONATE 21-ACETATE A solution of 100 mg. of the product of Example 25B in 2 ml. of anhydrous pyridine is treated with 100 mg. of p-toluenesulfonyl chloride and the mixture is stirred for 1 hour. The solution is poured into aqueous sulfuric acid and ice, and the resulting precipitate is removed by filtration, washed with water, dried, and crystallized from acetone-hexane to give 16u-methyl-6/3,17a,21-trihydroxy- 1,4 pregnadiene 3,11,20 trione 66 p-toluenesulfonate ZI-acetate.

I. 1GG-MECLHYLJ7a,21-DIHYDR0XY-1.5-PREGNADIENE 3,11,20-TRIONE 21-ACETATE The 6-p-toluenesulfonate 2l-acetate-1,4-pregnadiene of above Example 25H is reacted with zinc powder in the manner described in Example 138 to give 16u-methyl- 1711,21 dihydroxy 1,5 pregnadiene 3,11,20 trione 21-acetate.

The 21-aceta'te of the 16rx-methyl-l,S-pregnadiene prepared above is hydrolyzed to the corresponding 21- hydroxy compound in the manner described in Example 2.

EXAMPLE. 26

9oc-br0mo-16a-ethyl-1 701,21 -dihydr0xy-1 ,S-pregnadiene- 3,11,20-tri0ne A. 9a BROMO 16a. ETHYL 17a,21 DIHYDROXY 1,4- PREGNADIENE-3,11,20 lRIONE 2l-ACETATE The requisite intermediate, 9ec-bromo-16a-ethyl-115,1711, 21-trihydroxy-1,4-pregnadiene-3,20-dione 21-acetatc, is prepared as described in co-pending application Serial No. 733,843 of Rausser et al., filed May 8, 1958.

To a solution of 0.3 g. of 9a-bromo-16a-ethyl-1704,21- dihydroxy-l,5-pregnadiene-3,20-dione 2l-a'cetate in 15 ml. of acetic acid, there is added dropwise a solution of 60 mg. of chromium trioxide in 1 ml. of water and 3 ml. of acetic acid. The resultant mixture is allowed to stand for S'hours, then is diluted with water, and extracted with 17 methylene chloride. The organic extracts are washed with water, dried over magnesium sulfate, filtered and evaporated to a residue which is crystallized from methanol to give 9a-bromo-16a-ethyl-17a,21-dihydroxy-1,4-

wherein X is a halogen of atomic weight less than 126, Z is a lower alkyl, and R is lower alkanoyl.

3. fip-bromo-lA-pregnadienes having the formula:'

pregnadiene-3,l1,20-trione Zl-acetate. B. 65,9a DIBROMO 16a. E'IHYL 17a,21 DIHYDROXY- 4 1,'4-PREGNADIENE3,11,20-TRIONE 2l-ACETATE OH IOH The 9a-bromo-l6a-ethyl-1,4-pregnadiene of Example X A -wz 26A is brominated and the resultant product isolated in 10 J the manner described in Example 1A to give 613,90z-di- H3O bromo l(Set-ethyl-17a,2l-dihydroxy-1,4-pregnadiene-3,11,

20-trione ZI-acetate.

C. 9a, BROMO 16a ETHYL 1711,21 DIHYDROXY 1,5 O

PRAGNADIENE-3,ll,ZO-TRIONE 2l-ACETATE 6,8,9a dibromo 1Ger-ethyl-l7u,21-dihydroxy-1,4-pregnadiene-3,ll,20-trione 2l-acetate (500 mg.) is dissolved in 10 ml. of tetrahydrofuran, and 4 g. of magnesium turnings added. This mixture is stirred at room temperature Br wherein X is a halogen of atomic weight less than 126, Z is lower alkyl, and R is lower alkanoyl.

4. 6fi-bromo-1,4-pregnadienes of the formula:

for one hour, then 25 ml. of 5% aqueous ammonium sul- CH;

fate solution is added. This mixture is stirred another hour, then chloroform is added and the organic solvent (i=0 layer is separated from the aqueous layer. The chloro- 0 L--QH form solution is concentrated in vacuo to yield 9a-bromo- W W z 160: ethyl 17a,2l-dihydroxy-1,5-pregnadiene-3,l1,20-

trione ZI-acetate. CH3

D. 9a BROMO 16a ETHYL 1741,21 DIHYDROXY 1,5 PREGNADIENE-3,11,20-TRIONE The 21-acetate of Example 26C is hydrolyzed to the 30 corresponding 2l-alcohol with the aid of a culture of Flavobacterium dehydrogenans to yield 9u-bromo-16aethyl- 17a,2 l-dihydroxy-l ,5-pregnadiene-3, 1 1,20-trione.

Br wherein Z is lower alkyl and R is lower alkanoyl.

We claim: 5. Gfi-bromo-1,4-pregnadienes of the formula: 1. s-bromo-lA-pregnadienes of the following formula: CH;

CHzOR CH3 l wherein Z is lower alkyl, and R is lower alkanoyl.

6. 65 bromo 9a-fluoro-16-methyl-17a,21-dihydroxy- 1,4-pregnadiene3,11,20-tri0ne 21-acetate.

7. 6,3 brorno-l6-methyl-17a,21-dihydroxy-l,4-pregnadiene-3,11,20'-trione 21-acetate.

wherein X is a member of the group consisting of H and a halogen haw'ng an atomic weight less than 126; Y is 5 a member of the group consisting of O and (H, flOH); Z is a lower alkyl radical; and R is lower alkanoyl. 6/9 bromo 9 .fl r 16 thy1.11B 17 21-trihy- 2. 6fi-bromo-1,4-pregnadienes having the formula: 4 330 1m gp m 0H3 9 6,8-bromo-16-methyl-l1,8,l7a,2l-trihydroxy-1,4-preg- $13203 nad1ene-3,20-d1one 21-acetate. 0:0

0 OH M2 E30 References Cited in the file of this patent UNITED STATES PATENTS 2,816,902 Gould et a1. Dec. 17, 1957 2,887,499 Carvajal May 19, 1959 2,908,696 Nussbaum et al Oct. 13, 1959 OTHER REFERENCES 5 Arth et al.: 80, I.A.C.S., 3161-62 (1958). Br Oliveto et al.: 80, J.A.C.S., 4428 (1958). 

1. 6B-BROMO-1,4-PREGNADIENES OF THE FOLLOWING FORMULA: 